scholarly journals Structure of myosin filaments from relaxed Lethocerus flight muscle by cryo-EM at 6 Å resolution

2016 ◽  
Vol 2 (9) ◽  
pp. e1600058 ◽  
Author(s):  
Zhongjun Hu ◽  
Dianne W. Taylor ◽  
Michael K. Reedy ◽  
Robert J. Edwards ◽  
Kenneth A. Taylor
Keyword(s):  
Flight Muscle ◽  
Three Dimensional ◽  
Coiled Coil ◽  
Thick Filament ◽  
Coiled Coils ◽  
Muscle Physiology ◽  
Layer Arrangement ◽  
Thick Filaments ◽  
Unusual Position ◽  
Filament Structure

We describe a cryo–electron microscopy three-dimensional image reconstruction of relaxed myosin II–containing thick filaments from the flight muscle of the giant water bug Lethocerus indicus. The relaxed thick filament structure is a key element of muscle physiology because it facilitates the reextension process following contraction. Conversely, the myosin heads must disrupt their relaxed arrangement to drive contraction. Previous models predicted that Lethocerus myosin was unique in having an intermolecular head-head interaction, as opposed to the intramolecular head-head interaction observed in all other species. In contrast to the predicted model, we find an intramolecular head-head interaction, which is similar to that of other thick filaments but oriented in a distinctly different way. The arrangement of myosin’s long α-helical coiled-coil rod domain has been hypothesized as either curved layers or helical subfilaments. Our reconstruction is the first report having sufficient resolution to track the rod α helices in their native environment at resolutions ~5.5 Å, and it shows that the layer arrangement is correct for Lethocerus. Threading separate paths through the forest of myosin coiled coils are four nonmyosin peptides. We suggest that the unusual position of the heads and the rod arrangement separated by nonmyosin peptides are adaptations for mechanical signal transduction whereby applied tension disrupts the myosin heads as a component of stretch activation.

scholarly journals CryoEM Structure of Drosophila Flight Muscle Thick Filaments at 7Å Resolution

2020 ◽  
Author(s):  
Nadia Daneshparvar ◽  
Dianne W. Taylor ◽  
Thomas S. O’Leary ◽  
Hamidreza Rahmani ◽  
Fatemeh Abbasi Yeganeh ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Striated Muscle ◽  
Flight Muscle ◽  
Coiled Coil ◽  
Thick Filament ◽  
Fruit Fly ◽  
Actin Binding ◽  
Thick Filaments ◽  
Flight Muscles ◽  
Lethocerus Indicus

AbstractStriated muscle thick filaments are composed of myosin II and several non-myosin proteins. Myosin II’s long α-helical coiled-coil tail forms the dense protein backbone of filaments while its N-terminal globular head containing the catalytic and actin binding activities extends outward from the backbone. Here we report the structure of thick filaments of the flight muscle of the fruit fly Drosophila melanogaster at 7 Å resolution. Its myosin tails are arranged in curved molecular crystalline layers identical to flight muscles of the giant waterbug Lethocerus indicus. Four non-myosin densities are observed, three of which correspond to ones found in Lethocerus; one new density, possibly stretchin-Mlck, is found on the backbone outer surface. Surprisingly, the myosin heads are disordered rather than ordered along the filament backbone. Our results show striking myosin tail similarity within flight muscle filaments of two insect orders separated by several hundred million years of evolution.Significance StatementMyosin thick filaments are one of striated muscle’s key structures, but also one of its least understood. A key question is how the myosin a-helical coiled-coil tail is arranged in the backbone. At 7Å resolution, sufficient to resolve individual a-helices, the myosin tail arrangement in thick filaments from the flight muscle of the fruit fly Drosophila melanogaster is strikingly similar to the myosin tail arrangement in flight muscles of the giant waterbug Lethocerus indicus. Nearly every other thick filament feature is different. Drosophila and Lethocerus evolved separately >245 million years ago suggesting myosin tail packing into curved molecular crystalline layers forms a highly conserved thick filament building block and different properties are obtained by alterations in non-myosin proteins.

The relaxed crossbridge pattern in isolated rabbit psoas muscle thick filaments

1993 ◽  
Vol 105 (3) ◽  
pp. 841-848
Author(s):  
R.W. Kensler ◽  
M. Stewart
Keyword(s):  
Structural Difference ◽  
Psoas Muscle ◽  
Thick Filament ◽  
Rabbit Muscle ◽  
Fish Muscles ◽  
Thick Filaments ◽  
Filament Structure ◽  
The Individual ◽  
High Degree ◽  
Degree Of Similarity

Rabbit muscle is a major source of material for biochemical experiments and spin labelling studies of contraction, and so it is important to establish how closely this material resembles the frog and fish muscles usually used for structural studies. Previous studies have shown that relaxed rabbit muscle thick filaments lose the characteristic order of their crossbridges when they are cooled below about 15–19 degrees C, whereas the order of fish and frog muscles is retained above 0 degrees C. The lack of order has frustrated attempts to examine rabbit thick filament structure and has raised questions about how closely they might resemble other thick filaments. We have therefore developed a procedure for preserving the crossbridge order in isolated filaments. Electron microscopy of these thick filaments after either negative staining or metal shadowing has shown that the crossbridge pattern has a 43 nm axial repeat and is based on three near-helical strands. Computed transforms of either type of image show a series of layer lines confirming that the native relaxed pattern has been preserved, and computer reconstructions show the individual crossbridges lying on a slightly perturbed 3-stranded lattice. These data indicate an unexpectedly high degree of similarity between the rabbit and frog patterns and indicate that, in fully preserved material, there is little structural difference between the two thick filaments at the temperature at which each normally functions.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Molecular and ultrastructural defects in a Drosophila myosin heavy chain mutant: differential effects on muscle function produced by similar thick filament abnormalities.

1988 ◽  
Vol 107 (6) ◽  
pp. 2601-2612 ◽  
Author(s):  
P T O'Donnell ◽  
S I Bernstein
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Muscle Function ◽  
Flight Muscle ◽  
Thick Filament ◽  
Differential Sensitivity ◽  
Indirect Flight Muscle ◽  
Molecular Defect ◽  
Thick Filaments ◽  
Nuclease Mapping

We have determined the molecular defect of the Drosophila melanogaster myosin heavy chain (MHC) mutation Mhc and the mutation's effect on indirect flight muscle, jump muscle, and larval intersegmental muscle. We show that the Mhc1 mutation is essentially a null allele which results in the dominant-flightless and recessive-lethal phenotypes associated with this mutant (Mogami, K., P. T. O'Donnell, S. I. Bernstein, T. R. F. Wright, C. P. Emerson, Jr. 1986. Proc. Natl. Acad. Sci. USA. 83:1393-1397). The mutation is a 101-bp deletion in the MHC gene which removes most of exon 5 and the intron that precedes it. S1 nuclease mapping indicates that mutant transcripts follow two alternative processing pathways. Both pathways result in the production of mature transcripts with altered reading frames, apparently yielding unstable, truncated MHC proteins. Interestingly, the preferred splicing pathway uses the more distal of two available splice donor sites. We present the first ultrastrutural characterization of a completely MHC-null muscle and show that it lacks any discernable thick filaments. Sarcomeres in these muscles are completely disorganized suggesting that thick filaments play a critical role in sarcomere assembly. To understand why the Mhc1 mutation severely disrupts indirect flight muscle and jump muscle function in heterozygotes, but does not seriously affect the function of other muscle types, we examined the muscle ultrastructure of Mhc1/+ heterozygotes. We find that these organisms have a nearly 50% reduction in the number of thick filaments in indirect flight muscle, jump muscle, and larval intersegmental muscle. In addition, aberrantly shaped thick filaments are common in the jump muscle and larval intersegmental muscle. We suggest that the differential sensitivity of muscle function to the Mhc1 mutation is a consequence of the unique myofilament arrays in each of these muscles. The highly variable myofilament array of larval intersegmental muscle makes its function relatively insensitive to changes in thick filament number and morphology. Conversely, the rigid double hexagonal lattice of the indirect flight muscle, and the organized lattice of the jump muscle cannot be perturbed without interfering with the specialized and evolutionarily more complex functions they perform.

scholarly journals Hydrophobicity variations along the surface of the coiled-coil rod may mediate striated muscle myosin assembly in Caenorhabditis elegans.

1996 ◽  
Vol 135 (2) ◽  
pp. 371-382 ◽  
Author(s):  
P E Hoppe ◽  
R H Waterston
Keyword(s):  
Caenorhabditis Elegans ◽  
Striated Muscle ◽  
Coiled Coil ◽  
Thick Filament ◽  
Surface Hydrophobicity ◽  
Filament Assembly ◽  
Thick Filaments ◽  
Characteristic Variation ◽  
Muscle Myosin

Caenorhabditis elegans body wall muscle contains two isoforms of myosin heavy chain, MHC A and MHC B, that differ in their ability to initiate thick filament assembly. Whereas mutant animals that lack the major isoform, MHC B, have fewer thick filaments, mutant animals that lack the minor isoform, MHC A, contain no normal thick filaments. MHC A, but not MHC B, is present at the center of the bipolar thick filament where initiation of assembly is thought to occur (Miller, D.M.,I. Ortiz, G.C. Berliner, and H.F. Epstein. 1983. Cell. 34:477-490). We mapped the sequences that confer A-specific function by constructing chimeric myosins and testing them in vivo. We have identified two distinct regions of the MHC A rod that are sufficient in chimeric myosins for filament initiation function. Within these regions, MHC A displays a more hydrophobic rod surface, making it more similar to paramyosin, which forms the thick filament core. We propose that these regions play an important role in filament initiation, perhaps mediating close contacts between MHC A and paramyosin in an antiparallel arrangement at the filament center. Furthermore, our analysis revealed that all striated muscle myosins show a characteristic variation in surface hydrophobicity along the length of the rod that may play an important role in driving assembly and determining the stagger at which dimers associate.

scholarly journals A Structural Model for Phosphorylation Control of Dictyostelium Myosin II Thick Filament Assembly

1999 ◽  
Vol 147 (5) ◽  
pp. 1039-1048 ◽  
Author(s):  
Wenchuan Liang ◽  
Hans M. Warrick ◽  
James A. Spudich
Keyword(s):  
Structural Model ◽  
Myosin Ii ◽  
Coiled Coil ◽  
Thick Filament ◽  
Tail Region ◽  
Filament Assembly ◽  
Dictyostelium Myosin ◽  
Thick Filaments ◽  
Coil Structure

Myosin II thick filament assembly in Dictyostelium is regulated by phosphorylation at three threonines in the tail region of the molecule. Converting these three threonines to aspartates (3×Asp myosin II), which mimics the phosphorylated state, inhibits filament assembly in vitro, and 3×Asp myosin II fails to rescue myosin II–null phenotypes. Here we report a suppressor screen of Dictyostelium myosin II–null cells containing 3×Asp myosin II, which reveals a 21-kD region in the tail that is critical for the phosphorylation control. These data, combined with new structural evidence from electron microscopy and sequence analyses, provide evidence that thick filament assembly control involves the folding of myosin II into a bent monomer, which is unable to incorporate into thick filaments. The data are consistent with a structural model for the bent monomer in which two specific regions of the tail interact to form an antiparallel tetrameric coiled–coil structure.

scholarly journals THE FILAMENT LATTICE OF COCKROACH THORACIC MUSCLE

1968 ◽  
Vol 36 (3) ◽  
pp. 433-442 ◽  
Author(s):  
Martin Hagopian ◽  
David Spiro
Keyword(s):  
Fine Structure ◽  
Electron Microscope ◽  
Flight Muscle ◽  
Insect Flight ◽  
Thick Filament ◽  
Leucophaea Maderae ◽  
Thick Filaments ◽  
Femoral Muscle ◽  
Flight Muscles ◽  
Characteristic 3

The fine structure of the tergo-coxal muscle of the cockroach, Leucophaea maderae, has been studied with the electron microscope. This muscle differs from some other types of insect flight muscles inasmuch as the ratio of thin to thick filaments is 4 instead of the characteristic 3. The cockroach flight muscle also differs from the cockroach femoral muscle in thin to thick filament ratios and diameters and in lengths of thick filaments. A comparison of these latter three parameters in a number of vertebrate and invertebrate muscles suggests in general that the diameters and lengths of the thick filaments and thin to thick filament ratios are related.

scholarly journals Tomographic Three-dimensional Reconstruction of Insect Flight Muscle Partially Relaxed by AMPPNP and Ethylene Glycol

1997 ◽  
Vol 139 (3) ◽  
pp. 695-707 ◽  
Author(s):  
Holger Schmitz ◽  
Mary C. Reedy ◽  
Michael K. Reedy ◽  
Richard T. Tregear ◽  
Kenneth A. Taylor
Keyword(s):  
Ethylene Glycol ◽  
Flight Muscle ◽  
Insect Flight ◽  
Three Dimensional ◽  
Thick Filament ◽  
Power Stroke ◽  
Insect Flight Muscle ◽  
Target Zone ◽  
Dimensional Reconstruction ◽  
Myosin Subfragment 1

Rigor insect flight muscle (IFM) can be relaxed without ATP by increasing ethylene glycol concentration in the presence of adenosine 5′-[β′γ- imido]triphosphate (AMPPNP). Fibers poised at a critical glycol concentration retain rigor stiffness but support no sustained tension (“glycol-stiff state”). This suggests that many crossbridges are weakly attached to actin, possibly at the beginning of the power stroke. Unaveraged three-dimensional tomograms of “glycol-stiff” sarcomeres show crossbridges large enough to contain only a single myosin head, originating from dense collars every 14.5 nm. Crossbridges with an average 90° axial angle contact actin midway between troponin subunits, which identifies the actin azimuth in each 38.7-nm period, in the same region as the actin target zone of the 45° angled rigor lead bridges. These 90° “target zone” bridges originate from the thick filament and approach actin at azimuthal angles similar to rigor lead bridges. Another class of glycol-PNP crossbridge binds outside the rigor actin target zone. These “nontarget zone” bridges display irregular forms and vary widely in axial and azimuthal attachment angles. Fitting the acto-myosin subfragment 1 atomic structure into the tomogram reveals that 90° target zone bridges share with rigor a similar contact interface with actin, while nontarget crossbridges have variable contact interfaces. This suggests that target zone bridges interact specifically with actin, while nontarget zone bridges may not. Target zone bridges constitute only ∼25% of the myosin heads, implying that both specific and nonspecific attachments contribute to the high stiffness. The 90° target zone bridges may represent a preforce attachment that produces force by rotation of the motor domain over actin, possibly independent of the regulatory domain movements.

scholarly journals Dependence of thick filament structure in relaxed mammalian skeletal muscle on temperature and interfilament spacing

2021 ◽  
Vol 153 (3) ◽  
Author(s):  
Marco Caremani ◽  
Luca Fusi ◽  
Marco Linari ◽  
Massimo Reconditi ◽  
Gabriella Piazzesi ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Thick Filament ◽  
Structural Basis ◽  
Mammalian Skeletal Muscle ◽  
X Ray ◽  
Physiological Temperature ◽  
Thick Filaments ◽  
Filament Structure ◽  
Intact Muscle ◽  
Resting Muscle

Contraction of skeletal muscle is regulated by structural changes in both actin-containing thin filaments and myosin-containing thick filaments, but myosin-based regulation is unlikely to be preserved after thick filament isolation, and its structural basis remains poorly characterized. Here, we describe the periodic features of the thick filament structure in situ by high-resolution small-angle x-ray diffraction and interference. We used both relaxed demembranated fibers and resting intact muscle preparations to assess whether thick filament regulation is preserved in demembranated fibers, which have been widely used for previous studies. We show that the thick filaments in both preparations exhibit two closely spaced axial periodicities, 43.1 nm and 45.5 nm, at near-physiological temperature. The shorter periodicity matches that of the myosin helix, and x-ray interference between the two arrays of myosin in the bipolar filament shows that all zones of the filament follow this periodicity. The 45.5-nm repeat has no helical component and originates from myosin layers closer to the filament midpoint associated with the titin super-repeat in that region. Cooling relaxed or resting muscle, which partially mimics the effects of calcium activation on thick filament structure, disrupts the helical order of the myosin motors, and they move out from the filament backbone. Compression of the filament lattice of demembranated fibers by 5% Dextran, which restores interfilament spacing to that in intact muscle, stabilizes the higher-temperature structure. The axial periodicity of the filament backbone increases on cooling, but in lattice-compressed fibers the periodicity of the myosin heads does not follow the extension of the backbone. Thick filament structure in lattice-compressed demembranated fibers at near-physiological temperature is similar to that in intact resting muscle, suggesting that the native structure of the thick filament is largely preserved after demembranation in these conditions, although not in the conditions used for most previous studies with this preparation.

scholarly journals CryoEM structure of Drosophila flight muscle thick filaments at 7 Å resolution

2020 ◽  
Vol 3 (8) ◽  
pp. e202000823
Author(s):  
Nadia Daneshparvar ◽  
Dianne W Taylor ◽  
Thomas S O’Leary ◽  
Hamidreza Rahmani ◽  
Fatemeh Abbasiyeganeh ◽  
...  
Keyword(s):  
Striated Muscle ◽  
Flight Muscle ◽  
Myosin Ii ◽  
Coiled Coil ◽  
Fruit Fly ◽  
Actin Binding ◽  
Thick Filaments ◽  
Flight Muscles ◽  
Lethocerus Indicus ◽  
Globular Head

Striated muscle thick filaments are composed of myosin II and several non-myosin proteins. Myosin II’s long α-helical coiled-coil tail forms the dense protein backbone of filaments, whereas its N-terminal globular head containing the catalytic and actin-binding activities extends outward from the backbone. Here, we report the structure of thick filaments of the flight muscle of the fruit fly Drosophila melanogaster at 7 Å resolution. Its myosin tails are arranged in curved molecular crystalline layers identical to flight muscles of the giant water bug Lethocerus indicus. Four non-myosin densities are observed, three of which correspond to ones found in Lethocerus; one new density, possibly stretchin-mlck, is found on the backbone outer surface. Surprisingly, the myosin heads are disordered rather than ordered along the filament backbone. Our results show striking myosin tail similarity within flight muscle filaments of two insect orders separated by several hundred million years of evolution.

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